This proposal was designed to apply a genetic approach to construct novel combinations of the large and small subunits of ribulose 1,5-bisphosphate carboxylase-oxygenase (RuBPCase) and then to analyze its enzymatic properties. In the first year, we have successfully made the following interspecific hybrids. These are 1) Nicotiana sylvestris X N. tomentosiformis, 2) N. sylvestris X N. glauca, 3) N. sylvestris X N. tomentosiformis) X N. tomentosiformis, 4) (N. sylvestris X N. tomentosiformis) X N. glauca, 5) N. tabacum X N. langsdorffi, 6) N. tabacum X N. glutinosa. All hybrid plants contain RuBPCase of identical large subunits. However, the small subunit of each hybrid is different. They vary from one to four different polypeptides. In some cases, repeated backcross of several generations are required. The enzymes from the above hybrid plants have been crystallized. We are now in the process of identifying each enzyme by using isoelectric focusing and peptide mapping techniques. We have also investigated the effect of a nuclear mutation on the chemical and enzymatic properties of RuBPCase. Our results demonstrate that the yellow mutant plants have identical RuBPCase as that from the normal plants. The difference reported by other workers may be caused by the purification method used.